Nitric Oxide (NO) is a simple diatomic molecule that plays a diverse and complex role in cellular physiology. Less than 15 years ago NO was primarily considered a smog component formed during the combustion of fossil fuels mixed with air. However, as a result of the pioneering work of Ferid Murad et al. it is now known that NO is a powerful signaling compound and cytotoxic/cytostatic agent found in nearly every tissue including endothelial cells, neural cells and macrophages. Mammalian cells synthesize NO using a two step enzymatic process that oxidizes L-arginine to N-ω-hydroxy-L-arginine, which is then converted into L-citrulline and an uncharged NO free radical. Three different nitric oxide synthase enzymes regulate NO production. Neuronal nitric oxide synthase (NOS1, or nNOS) is formed within neuronal tissue and plays an essential role in neurotransmission; endothelial nitric oxide synthase (NOS3 or eNOS), is secreted by endothelial cells and induces vasodilatation; inducible nitric oxide synthase (NOS2 or iNOS) is principally found in macrophages, hepatocytes and chondrocytes and is associated with immune cytotoxicity.
Neuronal NOS and eNOS are constitutive enzymes that regulate the rapid, short-term release of small amounts of NO. In these minute amounts NO activates guanylate cyclase which elevates cyclic guanosine monophosphate (cGMP) concentrations which in turn increase intracellular Ca+2 levels. Increased intracellular Ca+2 concentrations result in smooth muscle relaxation which accounts for NO's vasodilating effects. Inducible NOS is responsible for the sustained release of larger amounts of NO and is activated by extracellular factors including endotoxins and cytokines. These higher NO levels are play a key role in cellular immunity.
Medical research is rapidly discovering therapeutic applications for NO including the fields of vascular surgery and intervention cardiology. Procedures used to clear blocked arteries such as percutaneous transluminal coronary angioplasty (PTCA) (also known as balloon angioplasty) and atherectomy and/or stent placement can result in vessel wall injury at the site of balloon expansion or stent deployment. In response to this injury a complex multi-factorial process known as restenosis can occur whereby the previously opened vessel lumen narrows and becomes re-occluded. Restenosis is initiated when thrombocytes (platelets) migrating to the injury site release mitogens into the injured endothelium. Thrombocytes begin to aggregate and adhere to the injury site initiating thrombogenesis, or clot formation. As a result, the previously opened lumen begins to narrow as thrombocytes and fibrin collect on the vessel wall. In a more frequently encountered mechanism of restenosis, the mitogens secreted by activated thrombocyles adhering to the vessel wall stimulate overproliferation of vascular smooth muscle cells during the healing process, restricting or occluding of the injured vessel lumen. The resulting neointimal hyperplasia is the major cause of a stent restenosis.
Recently, NO has been shown to significantly reduce thrombocyte aggregation and adhesion; this combined with NO's directly cytotoxic/cytostatic properties may significantly reduce vascular smooth muscle cell proliferation and help prevent restenosis. Thrombocyte aggregation occurs within minutes following the initial vascular insult and once the cascade of events leading to restenosis is initiated, irreparable damage can result. Moreover, the risk of thrombogenesis and restenosis persists until the endothelium lining the vessel lumen has been repaired. Therefore, it is essential that NO, or any anti-restenotic agent, reach the injury site immediately.
One approach for providing a therapeutic level of NO at an injury site is to increase systemic NO levels prophylacticly. This can be accomplished by stimulating endogenous NO production or using exogenous NO sources. Methods to regulate endogenous NO release have primarily focused on activation of synthetic pathways using excess amounts of NO precursors like L-arginine, or increasing expression of Nitric oxide synthase (NOS) using gene therapy. U.S. Pat. Nos. 5,945,452, 5,891,459 and 5,428,070 describe sustained NO elevation using orally administrated L-arginine and/or L-lysine. However, these methods have not been proven effective in preventing restenosis. Regulating endogenously expressed NO using gene therapy techniques remains highly experimental and has not yet proven safe and effective. U.S. Pat. Nos. 5,268,465, 5,468,630 and 5,658,565, describe various gene therapy approaches.
Exogenous NO sources such as pure NO gas are highly toxic, short lived and relatively insoluble in physiological fluids. Consequently, systemic exogenous NO delivery is generally accomplished using organic nitrate prodrugs such as nitroglycerin tablets, intravenous suspensions, sprays and transdermal patches. The human body rapidly converts nitroglycerin into NO; however, enzyme levels and co-factors required to activate the prodrug are rapidly depleted, resulting in drug tolerance. Moreover, systemic NO administration can have devastating side effects including hypotension and free radical cell damage. Therefore, using organic nitrate prodrugs to maintain systemic anti-restenotic therapeutic blood levels is not currently possible.
Therefore, considerable attention has been focused on localized, or site specific, NO delivery to ameliorate the disadvantages associated with systemic prophylaxis. Implantable medical devices and/or local gene therapy techniques including medical devices coated with NO-releasing compounds, or vectors that deliver NOS genes to target cells, have been evaluated. Like their systemic counterparts, gene therapy techniques for the localized NO delivery have not been proven safe and effective. There are still significant technical hurdles and safety concerns that must be overcome before site-specific NOS gene delivery will become a reality.
However, significant progress has been made in the field of localized exogenous NO application. As previously discussed, to be effective at preventing restenosis an inhibitory therapeutic such as NO must be administered for a sustained period at therapeutic levels. Consequently, any NO-releasing medical device used to treat restenosis must be suitable for long term or permanent implantation. An ideal candidate device is the arterial stent. Therefore, a stent that safely provides therapeutically effective amounts of NO to a precise location would represent a significant advance in restenosis prevention.
Nitric oxide-releasing compounds suitable for in vivo applications have been developed by a number of investigators. As early as 1960 it was demonstrated that nitric oxide gas could be reacted with amines to form NO-releasing anions having the following general formula (1):R—R′N—N(O)NO−  (1)wherein R and R′ are ethyl. Salts of these compounds could spontaneously decompose and release NO in solution. (R. S. Drago et al J. Am. Chem. Soc. 1960, 82, 96-98.)
Nitric oxide-releasing compounds with sufficient stability at body temperatures to be useful as therapeutics were ultimately developed by Keefer et al. as described in U.S. Pat. Nos. 4,954,526, 5,039,705, 5,155,137, 5,212,204, 5,250,550, 5,366,997, 5,405,919, 5,525,357 and 5,650,447 and in J. A. Hrabie et al, J. Org. Chem. 1993, 58, 1472-1476, all of which are herein incorporated by reference. Briefly, Hrabie et al. describes NO-releasing intramolecular salts (zwitterions) having the general formula (2):RN[N(O)NO]−(CH2)xNH2+R′  (2)The [N(O)NO]− (abbreviated herein after at NONO) containing compounds thus described release NO via a first order reaction that is predictable, easily quantified and controllable. This is in sharp contrast to other known NO-releasing compounds such as the S-nitrosothiol series as described in U.S. Pat. Nos. 5,380,758, 5,574,068 and 5,583,101.
Stable NO-releasing compounds of formula 2 have been coupled to amine containing polymers. U.S. Pat. No. 5,405,919 (“the '919 patent”) describes biologically acceptable polymers that may be coupled to NO-releasing groups including polyolefins, such as polystyrene, polypropylene, polyethylene, polyterafluoroethylene and polyvinylidene, and polyethylenimine, polyesters, polyethers, polyurethanes and the like. Medical devices, such as arterial stents, composed of these polymers represent a potential means for the site-specific delivery of NO. However, polymeric stents and other medical devices are not necessarily appropriate for all in situ applications where concentrated, localized NO delivery is desired. Many applications require rigid and semi-flexible metallic devices for optimum, long-term performance.
Expandable metal stents have proven to provide better support for arteries than their polymeric counterparts. However, unlike the polymers described above, biocompatible metals such as nickel, titanium, stainless steel and mixtures thereof, do not possess reactive surface amines. Therefore, attempts have been made to provide metallic medical devices with polymeric surfaces by coating them with amine containing polymers. However, many of these methods can be expensive and may diminish the safety and effectiveness of the medical devices, especially expandable stents. Stents coated with polymers such as those described in the '919 patent can be successfully treated to provide a sustained release of NO. However, there remains a risk that the polymer covering may fracture during stent expansion and deployment. Such fractures may result in fragments being released downstream, potentially blocking narrower arteriolae. Moreover, polymer coated metallic stents are more prone to thrombogenesis development than uncoated metallic stents. Consequently, polymer coated metallic medical devices that release NO have not been widely used.
Therefore, there is a need for NO-releasing metallic medical devices that do not rely on polymeric coatings. Consequently, it is an object of the present invention to provide metallic medical devices that deliver NO to specific anatomical sites within the mammalian body.
It is another object of the present invention to provide metallic medical devices capable of sustained NO release from their surfaces without the use of polymeric coatings.
It is yet another object of the present invention to provide metallic stents capable of the sustained release of NO from their surfaces in amounts sufficient to prevent restenosis.
It is yet another object of the present invention to provide methods for preventing occlusions in arteries, thus preventing restenosis.
It is another object of the present invention to provide methods for treating ischemic heart disease and simultaneously preventing restenosis by deploying an uncoated metal stent with a NO-releasing surface.